Strong expression of the calreticulin gene in the liver of Rana rugosa tadpoles, but not adult frogs.

Abstract

In the present paper we report the purification of calreticulin (CLT) from livers of the frog, Rana rugosa, the cloning and sequencing of its cDNA, and the CLT gene expression. CLT with M(r) = 52 kDa, estimated by SDS-PAGE, was purified from frog livers. Using rat CLT cDNA as a probe, a 2.4-kilobase frog cDNA clone was isolated from a frog liver cDNA library. The cDNA encoded 419 amino acids including an 18-residue NH2-terminal signal sequence that was 76% homologous to the rat CLT sequence and was 84% homologous to the partial sequence of Xenopus laevis CLT (Treves et al. [1992] Biochem. J. 287:579-581). Phylogenetic relationships estimated from the amino acid sequence of CLTs showed no pronounced variation between the two frog species, R. rugosa and X. laevis. Northern blot analysis indicated that the CLT mRNA level was very high in the liver of tadpoles, but extremely low in adult frogs. Expression levels were also very high in the premature ovary, while moderate expression was observed in the testis and brain of adult frogs. However, there was little histological change in the liver of tadpoles during development. Furthermore, CLT was recognized by Western blot analysis of total proteins in the liver of adult frogs. Immunostaining showed that CLT was distributed in the cytoplasm of liver cells. These results suggest that the expression of the CLT gene is tissue-dependent in the frog, R. rugosa, and that CLT probably functions biochemically in liver cells even when its gene expression is low.